The objectives of this research are to continue the purification, isolation and characterization of human melanoma-associated antigens (MAA), to study the mechanisms of their release by viable tumor cells. If successfully purified, MAA will be used to develop improved assays of humoral immunity to, and circulating antigens associated with, melanoma. The approach will be to radiolabel macromolecules on the surface of melanoma cells by the lactoperoxidase technique. Macromolecules will be solubilized by lysis in non-ionic detergents, or if "shed," collected in soluble form from culture media. MAA will be identified by immunoprecipitation with protein A-Sepharose and SDS-PAGE, utilizing polyclonal and monoclonal antimelanoma antibodies. Conventional biochemical procedures and solid phase immunoadsorbents will be used for radioimmunoassay and comparison with labeled macromolecules on control cells. Using these approaches, we have developed a polyvalent melanoma tumor antigen vaccine which contains a broad spectrum of MAAs defined by monoclonal and polyclonal melanoma antibodies and which is free of HLA-Dr antigens and FCS proteins. The immunogenicity and toxicity of this vaccine in humans is now being evaluated in a phase I trial. Several aspects of MAA biology relevant to understanding or modification of tumor growth will be studied, including: (1)\the immunogenicity of MAA in vivo and methods of enhancing it; (2)\the mechanisms of release of MAA by viable cells; and (3)\the effect of MAA release on the susceptibility of tumor cells to immune destruction.